Dieter Sell

Biotechnological production of 2-phenylethanol

Abstract. 2-Phenylethanol (2-PE) is an important flavour and fragrance compound with a rose-like odour. Most of the worlds annual production of several thousand tons is synthesised by chemical means but, due to increasing demand for natural flavours, alternative production methods are being sought. Harnessing the Ehrlich pathway of yeasts by bioconversion of L-phenylalanine to 2-PE could be an option, but in situ product removal is necessary due to product inhibition. This review describes the microbial production of 2-PE, and also summarizes the chemical syntheses and the market situation.

Methanol-based industrial biotechnology: current status and future perspectives of methylotrophic bacteria

Methanol is one of the building blocks in the chemical industry and can be synthesized either from petrochemical or renewable resources, such as biogas. Bioprocess technology with methylotrophic bacteria is well established, as illustrated by large-scale single-cell protein production in the past. During recent years, the first genomes of methylotrophs have been sequenced and significant progress in elucidating their metabolism has been made. In addition, the tool set for genetic engineering of methylotrophic bacteria has expanded greatly and strategies to produce fine and bulk chemicals with methylotrophs have been described. This review highlights the potential of these bacteria for the development of economically competitive bioprocesses based on methanol as an alternative carbon source, bringing together biological, technical and economic considerations.

Detection of the microbial activity of aerobic heterotrophic, anoxic heterotrophic and aerobic autotrophic activated sludge organisms with an electrochemical sensor

The microbial activity of aerobic heterotrophic, anoxic heterotrophic and aerobic autotrophic microorganisms in biological wastewater treatment was determined by means of an electrochemical bioactivity sensor. The development of the sensor resulted in a system which can determine the microbial activities that are relevant for effective wastewater treatment. The signals of the sensor system are proportional to the substrate degradation and it can show inhibiting effects on the biomass. The most important advantages of the system are: it is independent of O2 consumption, the three most important types of metabolic activities in wastewater technology can be measured with one sensor, furthermore the measurement is suitable for automation and it is on-line. The result is a potential for the optimization of processes based on microbial activity.

Regio- and Stereoselective Fungal Oxyfunctionalisation of Limonenes

Selective transformations of limonene by asco- and basidiomycetes were investigated. On the shake flask scale, Penicillium citrinum hydrated R-(+)-limonene to α-terpineol [83% regioselectivity (rs), more than 80 mg 1-1 product yield], and Gongronella butleri catalysed the terminal oxidation to yield perillyl alcohol (60% rs, 16 mg 1-1). On the laboratory bioreactor scale, Penicillium digitatum produced a peak concentration of 506 mg α-terpineol 1-1 in the fed-batch mode, equivalent to a theoretical yield of 67%, and no volatile by-products were found. Fusarium proliferatum transformed R-(+)-limonene enantiospecifically to cis-(+)- carveol (98.6% ee, more than 35 mg 1-1 product yield) and S-(-)-limonene predominantly to trans-(-)-carveol (96.3% ee). Pleurotus sapidus selectively dehydrogenised the accumulating trans-(-)-carveol to the corresponding enantiopure R-(-)-carvone. The results show that a careful selection of strain and bioprocess parameters may improve both the yield and the optical purity of a desired product.

Biotechnological production of terpenoid flavour and fragrance compounds in tailored bioprocesses

Abstract Integrated bioprocess concepts specific to the requirements of monoterpenes as precursors of microbial transformations were developed. R -Limonene and α-pinene were used as model precursors and bacteria and higher fungi served as biocatalysts. A closed-gas-loop bioreactor was set up to address the volatility of terpenoids. By this means α-terpineol was produced from R -limonene at the gram per litre scale with Penicillium digitatum. E. coli overexpressing a P450 BM-3 mutant served as biocatalyst to produce several hundreds of milligrams of pinene oxide, verbenol, and myrtenol from α-pinene. Here, an aqueous-organic two-phase bioreactor with diisononylphthalate as organic solvent was used for in situ precursor supply and product recovery.

CHARACTERIZATION OF STARTER CULTURES BY DETERMINING MICROBIAL ACTIVITY USING AN ELECTROCHEMICAL GEL CELL

ABSTRACT The application of starter cultures in the food industry is a widely used method of deliberately initiating fermentations of foodstuffs. Batch-dependent fluctuations in the activity of starter cultures can have an adverse effect on the quality of the products to be made using them. The amperometric sensor system presented in this study is able to detect deficiencies in the quality of starter cultures, thus enabling the end-user to adapt production steps according to the culture applied. The principle of the sensor system is current measurement by way of a three-electrode system. The current signal is based on a mediator, which has been reduced by microorganisms, immobilised in a gel together with the microorganisms.

Novel rapid in vitro cytotoxicity test on mammalian cells based on an electrochemical measuring method

Abstract The proliferation of pharmaceuticals and chemicals and the increasing manufacture of medical goods have made an assessment of their potential toxicological risks to man and the environment indispensable. Hence, a wide range of tests has been developed in order to generate data on the harmful effects of chemicals and pharmaceuticals on organisms. Initially, toxicological data were commonly collected from tests involving animals; however, on account of ethical objections, they are gradually being replaced by in vitro cytotoxicity tests. In this work, a new in vitro screening method for the determination of cytotoxicity, based on a novel electrochemical bioactivity sensor system, was implemented. The evaluation was based on current–time curves of a potentiostatic measurement proportional to the reduction of mediator molecules by mammalian cells. Depending on the number of reduced mediator molecules used by the cells, a current signal is produced, which provides information about the viability of living cells. By adding a toxic test substance to the cell, a reduction in the current signal can be observed, depending on the cytotoxicity of the substance. It is possible to quickly create a specific cytotoxic curve and to determine the corresponding inhibitory concentrations (IC50 value). First tests were performed on three mammalian cell lines and eight model compounds using this electrochemical measurement system. The IC50 values obtained corresponded well with the toxicity determined with an established reference cytotoxicity assay (MTT test) and with data reported in the literature.

Einsatz eines Sensors zurOnline-Bestimmung der mikrobiellen Aktivität

ZusammenfassungMit einem Bioaktivitätssensor kann die Aktivität der Mikroorganismen im Belebtschlamm gemessen werden. Das Signal des Sensors reagiert empfindlich auf eine Veränderung des Substratangebots und das Auftreten von toxischen Stoffen. Mit dem Bioaktivitätssensor ist es damit einerseits möglich,online die Belebungsstufe hinsichtlich ihrer mikrobiellen Aktivität zu kontrollieren und andererseits die zufließenden Abwässer bezüglich ihres toxischen Potentials zu überwachen.AbstractThe activity of microorganisms in the activated sludge can be measured with a bio-activity sensor. The signal of the sensor reacts sensitively to changes in substrate concentrations and the appearance of toxic materials. With the sensor, it is possible to check biological wastewater treatment plants online with regard to their microbial activity and to supervise the flowing sewage with reference to its toxic potential.

Lilac aldehydes and lilac alcohols as metabolic by-products of fungal linalool biotransformation

Nineteen different fungi were screened to test their capability to convert (±)-linalool into lilac aldehydes and lilac alcohols. Solid phase microextraction (SPME) in combination with GC-MS was used to identify the target compounds by comparing retention times and mass spectra with chemically synthesised standards. Most of the strains showed specific and complex product spectra, often with linalool oxides and dihydrolinalool as major metabolites. Aspergillus niger DSM 821, Botrytis cinerea 5901/02 and Botrytis cinerea 02/FB II/2.1 generated the desired lilac fragrance compounds as metabolic by-products.

Insights into the applicability of microbial fuel cells in wastewater treatment plants for a sustainable generation of electricity

ABSTRACT Microbial fuel cells (MFCs) are often discussed as a part of a sustainable generation of electricity for the coming ‘energy revolution’. In particular, the application of MFCs in wastewater treatment plants (WWTPs) are often regarded as an attractive alternative to reduce costs while generating electricity. Field surveys are necessary to show the applicability of MFCs in WWTPs considering daily fluctuations and environmental effects such as rain events affecting the MFC performance remarkably. In this study, a MFC system was tested in four municipal WWTPs using different modes of operation. A correlation between current densities and sludge loading (SL) was identified. At low SLs, the activated sludge needs a large amount of the energy derived from the substrate for the maintenance metabolism resulting in quite low current densities of the MFC. At high SLs much more of the energy can be transferred from the activated sludge to the electrode, resulting in higher currents. Furthermore, the effect of environmental conditions on the current densities was evaluated. WWTPs have daily fluctuations depending on the wastewater composition, weather phenomena and population equivalents. Our data show that these daily fluctuations can only be observed in the MFC performance at WWTPs below 50,000 population equivalents.